Dataset Information

Ex-Vivo Equine Cartilage Explant Osteoarthritis Model - A Metabolomics and Proteomics Study

ABSTRACT: Osteoarthritis (OA) is an age-related degenerative musculoskeletal disease characterised by loss of articular cartilage, synovitis, abnormal bone proliferation and subchondral bone sclerosis. The underlying pathogenesis of OA is yet to be fully elucidated with no OA specific biomarkers in clinical use. Nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry (MS) allow identification of the global metabolome and proteome respectively. During this study, ex-vivo equine cartilage explants (n=5) were incubated in TNF-α/IL-1β supplemented culture media for 8 days, with media removed and replaced at 2, 5 and 8 days. Acetonitrile metabolite extractions of 8 day cartilage explants and media samples at all time points underwent 1H NMR metabolic analysis with media samples also undergoing MS proteomic analysis. Within the cartilage, metabolites glucose and lysine were elevated following TNF-α/IL-1β treatment whilst adenosine, alanine, betaine, creatine, myo-inositol and uridine levels decreased. Within the culture media, four, four and six metabolites were identified as being differentially abundant between control and treatment groups for 1-2 day, 3-5 day and 6-8 day time points respectively. Culture media proteomics identified 154, 138 and 72 proteins differentially abundant, with > 2 fold change, between control and treatment groups for 1-2 day, 3-5 day and 6-8 day time points respectively. Nine potential novel OA neopeptides were elevated in treated media. This is the first study to use a multi ‘omics’ approach to simultaneously investigate the metabolomic profile of ex-vivo cartilage and metabolomic/proteomic profiles of culture media using the TNF-α/IL-1β ex-vivo OA cartilage model. This study has identified a panel of metabolites, proteins and extracellular matrix derived neopeptides which are differentially abundant during an early phase of the OA model which may provide further information on underlying disease pathogenesis, allow potential translation for clinical markers and possible novel therapeutic targets.

OTHER RELATED OMICS DATASETS IN: MTBLS1495

INSTRUMENT(S): Q Exactive

ORGANISM(S): Equus Caballus (horse)

TISSUE(S): Cell Culture

SUBMITTER: James Anderson

LAB HEAD: Mandy Jayne Peffers

PROVIDER: PXD017153 | Pride | 2020-08-03

REPOSITORIES: Pride

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Publications

Ex Vivo Equine Cartilage Explant Osteoarthritis Model: A Metabolomics and Proteomics Study.

Anderson James R JR Phelan Marie M MM Foddy Laura L Clegg Peter D PD Peffers Mandy J MJ

Journal of proteome research 20200806 9

Osteoarthritis is an age-related degenerative musculoskeletal disease characterized by loss of articular cartilage, synovitis, and subchondral bone sclerosis. Osteoarthritis pathogenesis is yet to be fully elucidated with no osteoarthritis-specific biomarkers in clinical use. Ex vivo equine cartilage explants (n = 5) were incubated in tumor necrosis factor-α (TNF-α)/interleukin-1β (IL-1β)-supplemented culture media for 8 days, with the media removed and replaced at 2, 5, and 8 days ...[more]

PMID: 32701294

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Project description:Autologous chondrocyte transplantation (ACT) is a routine technique to regenerate focal cartilage lesions. However, patients with osteoarthritis (OA) are lacking an appropriate long-lasting treatment alternative, partly since it is not known if chondrocytes from OA patients have the same chondrogenic differentiation potential as chondrocytes from donors not affected by OA. Articular chondrocytes from patients with OA undergoing total knee replacement (Mankin Score >3, AhlbÃ¤ck Score >2) and from patients undergoing ACT, here referred to as normal donors (ND), were isolated applying protocols used for ACT. Their chondrogenic differentiation potential was evaluated both in high-density pellet and scaffold (Hyaff-11) cultures by histological proteoglycan assessment (Bern Score) and immunohistochemistry for collagen types I and II. Chondrocytes cultured in monolayer and scaffolds were subjected to gene expression profiling using genome-wide oligonucleotide microarrays. Expression data were verified by using quantitative RT-PCR. Chondrocytes from ND and OA donors demonstrated accumulation of comparable amounts of cartilage matrix components, including sulphated proteoglycans and collagen types I and II. The mRNA expression of cartilage markers (COL2A1, COMP, aggrecan, CRTL1, SOX9) and genes involved in matrix synthesis (biglycan, COL9A2, COL11A1, TIMP4, CILP2) was highly induced in 3D cultures of chondrocytes from both donor groups. Genes associated with hypertrophic or OA cartilage (COL10A1, RUNX2, periostin, ALP, PTHR1, MMP13, COL1A1, COL3A1) were not significantly regulated between the two groups of donors. The expression of 661 genes, including COMP, FN1, and SOX9, were differentially regulated between OA and ND chondrocytes cultured in monolayer. During scaffold culture, the differences diminished between the OA and ND chondrocytes, and only 184 genes were differentially regulated. Only few genes were differentially expressed between OA and ND chondrocytes in Hyaff-11 culture. The risk of differentiation into hypertrophic cartilage does not seem to be increased for OA chondrocytes. Our findings suggest that the chondrogenic capacity is not significantly affected by OA and OA chondrocytes fulfill the requirements for matrix-associated ACT. Experiment Overall Design: Gene expression profiles of monolayer cultures (ML; passage 2) and Hyaff-11 scaffold cultures (3D; 14 days in vitro) of chondrocytes from 3 normal donors (ND; underwent ACT treatment) and 3 donors suffering from Osteoarthritis (OA; underwent knee replacement surgery) were determined. Comparative analyses between 3D and ML cultures (3D vs. ML) were performed to assess differentiation capacity of ND and OA chondrocytes. Furthermore, OA-related differences were determined comparing OA and ND monolayers as well as scaffold cultures (each OA vs. ND).

Dataset Information

Ex-Vivo Equine Cartilage Explant Osteoarthritis Model - A Metabolomics and Proteomics Study

Publications

<i>Ex Vivo</i> Equine Cartilage Explant Osteoarthritis Model: A Metabolomics and Proteomics Study.

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